Parotid acinar cells secrete proteins via two pathways that are distinct from salivary gland exocytosis: a constitutive-like pathway, insensitive to secretory stimulation and operating continuously, and a minor regulated pathway, which responds to low level stimulation. These two secretory pathways are hypothesized to provide the continual and neurally modulatable components of basal salivary secretion. The minor pathway has also been suggested to augment salivary output by priming secretory granule exocytosis. Long term goals of this project are to trace these pathways and understand their regulation and function at a molecular level using rat parotid glands as an experimental system. The first specific aim is to identify the carriers of the minor regulated pathway by a combination of techniques including biosynthetic labeling, subcellular fractionation, and immunofluorescence microscopy. The second specific aim is to develop a permeabilized cell system that faithfully reiterates secretion by the minor regulated pathway and by granule exocytosis. This system will be used to analyze the function of the minor regulated pathway and its role in priming for granule release. The final aim of this project addresses a novel aspect of stimulus-secretion coupling in salivary acinar cells. The goal is to determine if stimulus-evoked relocation of the protein SNAP-23 from the basolateral membrane to the apical secretory apparatus serves to link cellular stimulation to exocytosis, and if so, how. In this aim, relocation will be examined further using parotid lobules and permeabilized acinar cells in experiments that include immunofluorescence and immunoelectron microscopy, antibody perturbation, and analysis of molecular interactions. Oral/salivary disorders such as xerostomia and ductal obstruction are primary or secondary consequences of impaired salivary secretion that may emphasize dysfunction in both basal and stimulated secretion. The new directions being taken in specifically addressing basal secretion and in examining what is potentially a key event in stimulus-secretion coupling may help to uncover the defects that cause the salivary dysfunctions. The findings may be broadly applicable in exocrine secretory cells.